Protein-O-carboxylmethyltransferase (E.C. 2.1.1.24; PCM) has been purified from rat brain cytosol and human erythrocyte cytosol. These enzymes have been subjected to amino acid analysis which indicated that rat and human PCM slightly differed in amino acid composition. Several lectin-affinity chromatography columns were used to describe possible terminal sugars. Also, kinetic analyses were performed indicating that the Km of both human and rat PCM for AdoMet was similar (about 10.0 MuM), and that this value was not changed when either porcine skin gelatin or calmodulin was used as a methyl acceptor. PCM was inhibited by S-adenosylhomocysteine with a Ki of about 1.0 MuM. We have also observed that irradiation of purified human PCM with ultraviolet light causes a covalent binding of the methyl group from S-adenosylmethionine to the enzyme. This unusual covalent binding may allow further elucidation into the mechanism of methyl transfer of this enzyme.